For the concentration measurement of metal ions such as potassium ions, various methods have hitherto been employed, such as flame photometry, chemical analysis and ion-selective electrode method. However, the flame photometry requires complicated procedures and has a poor capacity for sample processing. The chemical analysis also requires complicated procedures and the reagents to be used therein are expensive, so that it is not actually used in the field of clinical examination. The ion-selective electrode method, although it can be conducted by relatively simple procedures, has the disadvantage that the occurrence of errors in the measurement may be caused by deterioration of the electrode.
In recent years, there are several reports on a method for measurement of potassium ion concentration according to an enzymatic technique. For example, a colorimetric method has been reported in which pyruvate oxidase is allowed to act on pyruvates which are produced by a reaction using pyruvate kinase and the resulting hydrogen peroxide is utilized for forming a quinoid dye under the coexistence of a chromogen and peroxidase, followed by a colorimetric analysis. However, this method is readily affected by endogenous pyruvates and pyruvate oxidase is only available at a high cost, thereby making it difficult to apply this method to commercial use.
In another method disclosed in Clin. Chem., vol. 35, No. 5, pp. 817-820, 1989, a decrease in absorbance is measured at the ultraviolet region (around 340 nm) of nicotinamide adenine dinucleotide of reduced type (NADH), and therefore, this method has the disadvantages that a decrease in the amount of NADH during the storage of reagents makes impossible the measurement of specimens having the possibility of exhibiting high measured values and that the improvement in sensitivity cannot be attained over a certain constant level because a decrease in absorbance is utilized therein. Moreover, as means for avoiding the influence of ammonium ions, glutamate dehydrogenase is usually used to eliminate these ammonium ions according to the scheme I as shown below. This results is another problem that the setting of absorbance or wavelength is difficult when ammonium ions of high concentration should be eliminated, because the wavelength range for measurement of potassium ion concentration overlaps with that for measurement of ammonium ion elimination and a great amount of co-enzymes is required to eliminate ammonium ions of high concentration ##STR1##
Further reported is a method for elimination of ammonium ions by use of glutamine synthetase and asparagine synthetase or by a combination of glutamate dehydrogenase and isocitrate dehydrogenase ("Rinsho-Kensa; Kiki-Shiyaku (Clinical Examination; Equipments and Reagents)", vol. 14, No. 2, April 1991). In the former method, however, glutamine or asparagine produced by the elimination reaction of ammonium ions has a tendency to give free ammonium ions. In the latter method, isocitric acid as a substrate of isocitrate dehydrogenase is too expensive to provide its reagent at a low cost when this method is applied to commercial use. Moreover, the addition of a metal chelating agent is required to terminate the reaction of isocitrate dehydrogenase, but when the concentration of metal ions is measured the addition of a metal chelating agent should be limited to the smallest possible extent because of its negative influence on the measurement. It is, therefore, difficult for these methods to find commercial applications to the measurement of metal ion concentration.